1. Biochemistry and Chemical Biology
  2. Cell Biology

eIF2B conformation and assembly state regulates the integrated stress response

  1. Michael Schoof
  2. Morgane Boone
  3. Lan Wang
  4. Rosalie Lawrence
  5. Adam Frost  Is a corresponding author
  6. Peter Walter  Is a corresponding author
  1. University of California, San Francisco, United States
  2. Howard Hughes Medical Institute, University of California, San Francisco, United States
https://doi.org/10.7554/eLife.65703
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  1. Michael Schoof
  2. Morgane Boone
  3. Lan Wang
  4. Rosalie Lawrence
  5. Adam Frost
  6. Peter Walter
(2021)
eIF2B conformation and assembly state regulates the integrated stress response
eLife 10:e65703.
https://doi.org/10.7554/eLife.65703
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Abstract

The integrated stress response (ISR) is activated by phosphorylation of the translation initiation factor eIF2 in response to various stress conditions. Phosphorylated eIF2 (eIF2-P) inhibits eIF2's nucleotide exchange factor eIF2B, a two-fold symmetric heterodecamer assembled from subcomplexes. Here, we monitor and manipulate eIF2B assembly in vitro and in vivo. In the absence of eIF2B's α-subunit, the ISR is induced because unassembled eIF2B tetramer subcomplexes accumulate in cells. Upon addition of the small-molecule ISR inhibitor ISRIB, eIF2B tetramers assemble into active octamers. Surprisingly, ISRIB inhibits the ISR even in the context of fully assembled eIF2B decamers, revealing allosteric communication between the physically distant eIF2, eIF2-P, and ISRIB binding sites. Cryo-EM structures suggest a rocking motion in eIF2B that couples these binding sites. eIF2-P binding converts eIF2B decamers into 'conjoined tetramers' with diminished substrate binding and enzymatic activity. Canonical eIF2-P-driven ISR activation thus arises due to this change in eIF2B's conformational state.

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All data generated or analysed during this study are included in the manuscript and supporting files.

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Author details

  1. Michael Schoof

    Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States
    Competing interests
    No competing interests declared.

  2. Morgane Boone

    Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7807-5542

  3. Lan Wang

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8931-7201

  4. Rosalie Lawrence

    Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States
    Competing interests
    No competing interests declared.

  5. Adam Frost

    Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States
    For correspondence
    adam.frost@ucsf.edu
    Competing interests
    Adam Frost, Reviewing editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2231-2577

  6. Peter Walter

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    For correspondence
    peter@walterlab.ucsf.edu
    Competing interests
    Peter Walter, PW is an inventor on U.S. Patent 9708247 held by the Regents of the University of California that describes ISRIB and its analogs. Rights to the invention have been licensed by UCSF to Calico..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6849-708X

Funding

Howard Hughes Medical Institute (Investigator Grant)

  • Peter Walter

Howard Hughes Medical Institute (HHMI Faculty Scholar Grant)

  • Adam Frost

Calico Life Sciences LLC

  • Peter Walter

The George and Judy Marcus Family Foundation

  • Peter Walter

Damon Runyon Cancer Research Foundation (Postdoctoral Fellowship)

  • Lan Wang

Jane Coffin Childs Memorial Fund for Medical Research (Postdoctoral Fellowship)

  • Rosalie Lawrence

Belgian-American Educational Foundation (Postdoctoral Fellowship)

  • Morgane Boone

Chan Zuckerberg Initiative (Investigator Grant)

  • Adam Frost

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2021, Schoof et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Michael Schoof
  2. Morgane Boone
  3. Lan Wang
  4. Rosalie Lawrence
  5. Adam Frost
  6. Peter Walter
(2021)
eIF2B conformation and assembly state regulates the integrated stress response
eLife 10:e65703.
https://doi.org/10.7554/eLife.65703

Share this article

https://doi.org/10.7554/eLife.65703

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Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics

    A point mutation in the nucleotide exchange factor eIF2B constitutively activates the integrated stress response by allosteric modulation

    Morgane Boone, Lan Wang ... Michael Schoof
    Research Advance Updated

    In eukaryotic cells, stressors reprogram the cellular proteome by activating the integrated stress response (ISR). In its canonical form, stress-sensing kinases phosphorylate the eukaryotic translation initiation factor eIF2 (eIF2-P), which ultimately leads to reduced levels of ternary complex required for initiation of mRNA translation. Previously we showed that translational control is primarily exerted through a conformational switch in eIF2’s nucleotide exchange factor, eIF2B, which shifts from its active A-State conformation to its inhibited I-State conformation upon eIF2-P binding, resulting in reduced nucleotide exchange on eIF2 (Schoof et al. 2021). Here, we show functionally and structurally how a single histidine to aspartate point mutation in eIF2B’s β subunit (H160D) mimics the effects of eIF2-P binding by promoting an I-State like conformation, resulting in eIF2-P independent activation of the ISR. These findings corroborate our previously proposed A/I-State model of allosteric ISR regulation.

    1. Biochemistry and Chemical Biology

    Disordered proteins interact with the chemical environment to tune their protective function during drying

    Shraddha KC, Kenny H Nguyen ... Thomas C Boothby
    Research Article

    The conformational ensemble and function of intrinsically disordered proteins (IDPs) are sensitive to their solution environment. The inherent malleability of disordered proteins, combined with the exposure of their residues, accounts for this sensitivity. One context in which IDPs play important roles that are concomitant with massive changes to the intracellular environment is during desiccation (extreme drying). The ability of organisms to survive desiccation has long been linked to the accumulation of high levels of cosolutes such as trehalose or sucrose as well as the enrichment of IDPs, such as late embryogenesis abundant (LEA) proteins or cytoplasmic abundant heat-soluble (CAHS) proteins. Despite knowing that IDPs play important roles and are co-enriched alongside endogenous, species-specific cosolutes during desiccation, little is known mechanistically about how IDP-cosolute interactions influence desiccation tolerance. Here, we test the notion that the protective function of desiccation-related IDPs is enhanced through conformational changes induced by endogenous cosolutes. We find that desiccation-related IDPs derived from four different organisms spanning two LEA protein families and the CAHS protein family synergize best with endogenous cosolutes during drying to promote desiccation protection. Yet the structural parameters of protective IDPs do not correlate with synergy for either CAHS or LEA proteins. We further demonstrate that for CAHS, but not LEA proteins, synergy is related to self-assembly and the formation of a gel. Our results suggest that functional synergy between IDPs and endogenous cosolutes is a convergent desiccation protection strategy seen among different IDP families and organisms, yet the mechanisms underlying this synergy differ between IDP families.